Quorum Sensing Regulation and Its Role In

Abraham Mikru PhD seminar AAU 2010

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Quorum Sensing Regulation And Its Role In The Legume-

Rhizobia Symbiosis

PhD Seminar By Abraham Mikru Teklemichael

Instructor: Fasil Assefa (PhD) Dpt. Biology, AAU 2010


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Contents

• Introduction• General principles of quorum sensing• The Lux paradigm• QS in some rhizobia• Response of host plant • Concluding remarks


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Introduction

• Microbes are not solitary

• Microbes interact with each other and their environment

• Microbial interaction is widespread & vast

• The focus here is on quorum sensing (QS) via acylhomoserine lactones (AHLs)


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What is QS?• Cell Density-dependent regulation of gene

expression mediated by diffusible signal molecules

• “Molecular dialogue”• Bacterial ‘talk’ to each other & ‘ccrosstalk’• Bacterial chemical communication or

signaling


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Why do bacteria communicate?

• Regulate gene expression concertedly• Orchestrate activities not achievable by a

single cell

• Coordinate population-wide behavioral response

• Behave as a multicellular organism• Rapid adaptation and survival


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Behaviors controlled by QS

Among others:• Bioluminescence• Virulence• Symbiosis• Biofilm • Horizontal gene transfer


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QS molecules (autoinducer)

• N-Acyl homoserine lactones (AHLs) or AI-1• In Gram negatives as well as rhizobia• Exclusively intraspecies communication

Hydrophilic head

Homoserine lactonering

Hydrophobic tailAcyl side chain

Variability•n= C4 -C18•R= H, O, OH•Unsaturation


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Synthesis and accumulation

SAM= S-adenosyl methionineACP= acyl-carrier protein

3. Accumulation depends on:

•Cell density•Biofilm•Microcolony•Microenvironment •Degraders

2. Movement across membrane

•Diffusion•Active/carrier mediated

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QS regulator (LuxR and homologues)

• AHL receptor and transcriptional regulator• Two regions of sequence conservation

Amino terminus binds to cognate AHLCarboxy terminus binds to target DNA

(Lux box)


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AHL degraders

• Unstable under alkaline pH• AHL degrading enzymes • Produced by several rhizobacteria• Potentially interfere with QS

Lactonase (1) and Acylase (3)


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When does QS occur?

At threshold [AHL] or when population is quorate

time

growth

Quorum sensing-regulated gene expression

Quorumsize


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How many cells in a Quorate population? Depends…

Increasing Cell Density

[AHLs]

Critical [AHL]Specific Genes ON

Low cell density High cell density


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General principle of QS and the Lux paradigm

• Cells make a signalling molecule (AHL) constitutively• The AHL diffuses freely out of the cell into the

surrounding medium• As the population of the cells grow, so does the

concentration of AHL in the growth medium• When the population (and hence AHL) concentration

reaches a threshold, the AHL moves back down its concentration gradient into the cell.


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Cont.

• The concentration of AHL inside the cell now increases and AHL can bind to luxR – the transcriptional activator protein

• The luxR-AHL conjugate activates gene expression

• This system is often referred to as “autoinduction” or

• Positive feedback loop control


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luxI codes for AHLSynthase

luxR codes forTranscriptional regulator

QS in Photobacterium fischeri (V.fischeri)


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Bioluminescence at high cell density in culture and light organ


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QS in Rhizobiacae

• QS now considered one of the requisite symbiotic signals

• Reported for many rhizobia

• In all cases AHLs are the signaling molecules

• The genI/genR (luxI/luxR homologues) system still remains


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QS in R. leguminosarum

Four QS systems known

• cinI/cinR• raiI/raiR• rhiI/rhiR• traI/traR


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cinI/cinR

• Located on chromosome

QS molecule is: • 3-OH-C14:1-HSL

Functions• Overall control of other QS systems• Stationary phase adaptation and survival• Growth inhibition (bacteriocin–like action)


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raiI/raiR

• Located on non-symbiotic plasmid

QS molecules:• C8-HSL, 3-OH-C8-HSL• Function: unknown


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rhiI/rhiR

• Located on symbiotic plasmid• Close to nod box

QS molecules• C6-HSL, C7-HSL and C8-HSL

Target genes• rhiI, rhiABC

Function• Influence nodulation


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traI/traR

• Located on symbiotic plasmidQS molecules• 3-oxo-C8-HSL• 3-OH-C14:1-HSL from recipient

Other regulatory genes• bisR, traM

Target genes and functions• Induce plasmid transfer operon


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QS network in R. leguminosarum


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traI/traR


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The R. etli Qs systems

• Possess orthologues of the cinI/cinR, raiI/raiR and traI/traR genes

• Target genes and functions are different• cinI/cinR: growth inhibition, symbiosome

development, nitrogen fixation and swarming motility

• raiI/raiR : Mutants show higher nodulation• traI/traR: conjugal plasmid (p42a)transfer


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Interaction of two QS in R. etli CNPAF 512


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S. meliloti QS systems

• sinI/sinR: EPS production, swarming motility and nodulation efficiency

• Involves several C12 to C18 AHLs • Some symbiotic genes dependent on sinR

but not on sinI. • What could be the signal molecules other

than SinI- made AHLS controls these genes?

• expR: role in EPSII synthesis


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S. meliloti (Rm1021 and Rm41)


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Mesorhizobium spp

Reported for several species

• M. huakuii: QS involved in biofilm formation• M. loti: three luxI-like genes influence

nodulation • M. tianshanense: QS involved in nodulation


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Rhizobium sp.NGR234

• traI/traR genes on symbiotic plasmid pNGR234a

• QS molecule is 3-oxo-C8-HSL

Functions• Conjugal plasmid transfer• Growth inhibition


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Bradyrhizobium japonicum

• QS inhibit nod gene expression• QS molecule is bradyoxetin not an AHL• Exhibit siderophor-like property• Maximal in iron depleted condition• AHLs detected in B. japonicum and B. elkani

but the target genes not known


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Bradyrhizobium japonicum


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Responses of host plants to QS

Plants ‘listen’ & respond to bacterial AHL QS• Medicago truncatula root respond to two

different AHLs by up regulating 154 proteins• The proteins function in defense,

transcriptional regulation, stress and plant hormone response

• Host plants also produce AHL-mimics potentially interfering with bacterial QS


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Concluding remarks

AHL QS plays several role in rhizobia-legume symbiosis

• EPS production,symbiosome development, Nodulation efficiency,Nitrogen fixation

• Plasmid transfer • Stationary phase adaptation and Growth

inhibition Host plants also “listen” and respond to

bacterial AHL-QS


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Thank You!


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Variations in structure of AHLs

A. tumefaciens, Rhizobium sp. NGR234, R. leguminosarum, B. S. meliloti, R. etli CFN42

R. leguminosarum, S. meliloti, R. etli CFN42

R. leguminosarum, S. meliloti

R. leguminosarum

S. meliloti

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